Layer-by-layer biofabrication of coronary covered stents with clickable elastin-like recombinamers

Producción CientíficaCoronary artery disease is the leading cause of death around the world. Endovascular stenting is the preferred treatment option to restore blood flow in the coronary arteries due to the lower perioperative morbidity when compared with more invasive treatment options. However, stent failure is still a major clinical problem, and further technological solutions are required to improve the performance of current stents. Here, we developed coronary stents covered with elastin-like recombinamers (ELRs) by exploiting a layer-by-layer technique combined with catalyst-free click chemistry. The resulting ELR-covered stents were intact after an in vitro simulated implantation procedure by balloon dilatation, which evidenced the elastic performance of the membrane. Additionally, the stents were mechanically stable under high flow conditions, which is in agreement with the covalent and stable nature of the click chemistry crosslinking strategy exploited during the ELR-membrane manufacturing and the successful embedding of the stent. Minimal platelet adhesion was detected after blood exposure in a Chandler loop as shown by scanning electron microscopy. The seeding of human endothelial progenitor cells (EPCs) on the ELR-membranes resulted in a confluent endothelial layer. These results prove the potential of this strategy to develop an advanced generation of coronary stents, with a stable and bioactive elastin-like membrane to exclude the atherosclerotic plaque from the blood stream or to seal coronary perforations and aneurysms, while providing a luminal surface with minimal platelet adhesion and favouring endothelialization.German federal and state governments (project StUpPD_330-18)Ministerio de Economía, Industria y Competitividad (projects PCIN-2015-010 / MAT2016-78903-R)Junta de Castilla y León (project VA317P18

Geometric bionics: Lotus effect helps polystyrene nanotube films get good blood compatibility

Various biomaterials have been widely used for manufacturing biomedical applications including artificial organs, medical devices and disposable clinical apparatus, such as vascular prostheses, blood pumps, artificial kidney, artificial hearts, dialyzers and plasma separators, which could be used in contact with blood^1^. However, the research tasks of improving hemocompatibility of biomaterials have been carrying out with the development of biomedical requirements^2^. Since the interactions that lead to surface-induced thrombosis occurring at the blood-biomaterial interface become a reason of familiar current complications with grafts therapy, improvement of the blood compatibility of artificial polymer surfaces is, therefore a major issue in biomaterials science^3^. After decades of focused research, various approaches of modifying biomaterial surfaces through chemical or biochemical methods to improve their hemocompatibility were obtained^1^. In this article, we report that polystyrene nanotube films with morphology similar to the papilla on lotus leaf can be used as blood-contacted biomaterials by virtue of Lotus effect^4^. Clearly, this idea, resulting from geometric bionics that mimicking the structure design of lotus leaf, is very novel technique for preparation of hemocompatible biomaterials

In vitro Removal of Therapeutic Drugs with a Novel Adsorbent System

Background/Aim: Substances in the middle molecular weight range have been shown to play a significant pathogenetic role in as diverse disorders as end-stage renal disease and multiple organ failure. To overcome the limitations in the amount removed by hemofilters, new sorbents with a high biocompatibility are actively being developed. Furthermore, biocompatible sorbents by their nonspecific adsorptive behavior could have great impact on detoxification treatment in exogenous intoxications. We performed an in vitro evaluation of a newly developed highly biocompatible sorbent cartridge (Betasorb(R)), examining its adsorptive capacity concerning therapeutic drugs. Methods: Uremic blood spiked with a range of therapeutic drugs was recirculated for 2 h in an in vitro hemoperfusion circuit containing a Betasorb device for hemoperfusion. The drug concentrations before and after the passage of the cartridge were measured, and the total amount removed was calculated. Results: The sorbent showed effective removal of glycopeptide antibiotics, digoxin, theophylline, phenobarbital, phenytoin, carbamazepine, and valproic acid. Moderate removal could be demonstrated for tacrolimus and cyclosporine A; aminoglycosides were removed to a small extent only. Conclusions: Betasorb hemoperfusion shows a potent adsorptive capacity concerning therapeutic drugs (except aminoglycosides) and could be of major value in the treatment of intoxications. On the other hand, drug monitoring and possible adjustments are necessary during Betasorb hemoperfusion to maintain the therapeutic ranges of the drugs in blood. Copyright (C) 2002 S. Karger AG, Basel

Interactions of siRNA loaded dextran nanogel with blood cells

Adsorption and internalization of siRNA loaded dextran nanogels by blood cells were determined using flow cytometry. Positively and negatively charged nanogels with various PEGylation degrees were compared in order to find a formulation showing minimal interactions with blood

Biocompatibility assessment of silk nanoparticles : hemocompatibility and internalization by human blood cells

Many nanoparticles are designed for use as potential nanomedicines for parenteral administration. However, emerging evidence suggests that hemocompatibility is important, but is highly particle- and test-bed dependent. Thus, knowledge of bulk material properties does not predict the hemocompatibility of uncharacterized nanoparticles, including silk nanoparticles. This study compares the hemocompatibility of silk versus silica nanoparticles, using whole human blood under quasi-static and flow conditions. Substantial hemocompatibility differences are noted for some nanoparticles in quasi-static versus dynamic studies; i.e., the inflammatory response to silk nanoparticles is significantly lower under flow versus quasi-static conditions. Silk nanoparticles also have very low coagulant properties - an observation that scales from the macro- to the nano-level. These nanoparticle hemocompatibility studies are complemented by preliminary live cell measurements to evaluate the endocytosis and trafficking of nanoparticles in human blood cells. Overall, this study demonstrates that nanoparticle hemocompatibility is affected by several factors, including the test bed design

Fabrication and properties of L-arginine-doped PCL electrospun composite scaffolds

The article describes fabrication and properties of composite fibrous scaffolds obtained by electrospinning of the solution of poly({\epsilon}-caprolactone) and arginine in common solvent. The influence of arginine content on structure, mechanical, surface and biological properties of the scaffolds was investigated. It was found that with an increase of arginine concentration diameter of the scaffold fibers was reduced, which was accompanied by an increase of scaffold strength and Young modulus. It was demonstrated that porosity and water contact angle of the scaffold are independent from arginine content. The best cell adhesion and viability was shown on scaffolds with arginine concentration from 0.5 to 1 % wt

Dynamic in vitro hemocompatibility testing – improving the signal to noise ratio

Assigning the hemocompatibility of small vascular implants is one of the great challenges in biomedical engineering. Due to the fact, that there are no widely approved test setups1, we decided to developed a modified CHANDLER-Loop system for dynamic in vitro hemocompatibility tests. The setup allows simultaneous testing of about 30 tube rings with an inner diameter of 2,4 mm. Cardiovascular implants can be placed into these tube rings. After filling them with anticoagulated blood and closing them to loops, they were placed on a rotating disc. While the loop is rotating, the blood remains in the lower part of the loop. The relative movement between the foreign surface and the blood sample induces a blood flow without the need of mechanical pumping. A first test series with the new test setup had been conducted to achieve a highly hemocompatible tube material. The results were compared to hemocompatibility tests done by LEMM, mentioned in the ISO 10993-42,3. We could prove, that Tygon S50HL® and Polyurethane achieve similar hemocompatibility values. In addition, we did in vitro tests of bare metal stents. We recorded a difference in the index of thrombocytes of about 17 % between the loops with three stents and the control. During further experiments the difference decreases to 8 %, which might be due to adhesion processes. The ability to measure these processes demonstrates the high sensitivity and very low background activation of the test setup. This can be assured by the recorded index of hemolysis, which had been less than 0,8 % in all our experiments. By developing and improving our modified CHANDLER-Loop system, we were able to establish a dynamic in vitro test setup for the hemocompatibility testing of small vascular implants. Because of its extremely low background activation and a high variability, the system serves as a benchmark for upcoming test setups.SFB-TR37 – Q2SFB-TR37 – B

Low molecular weight ϵ-caprolactone-pcoumaric acid copolymers as potential biomaterials for skin regeneration applications

ϵ-caprolactone-p-coumaric acid copolymers at different mole ratios (ϵ-caprolactone:p-coumaric acid 1:0, 10:1, 8:1, 6:1, 4:1, and 2:1) were synthesized by melt-polycondensation and using 4-dodecylbenzene sulfonic acid as catalyst. Chemical analysis by NMR and GPC showed that copolyesters were formed with decreasing molecular weight as p-coumaric acid content was increased. Physical characteristics, such as thermal and mechanical properties, as well as water uptake and water permeability, depended on the mole fraction of pcoumaric acid. The p-coumarate repetitive units increased the antioxidant capacity of the copolymers, showing antibacterial activity against the common pathogen Escherichia coli. In addition, all the synthesized copolyesters, except the one with the highest concentration of the phenolic acid, were cytocompatible and hemocompatible, thus becoming potentially useful for skin regeneration applications

Immuno Magnetic Thermosensitive Liposomes For Cancer Therapy

The present work describes the encapsulation of the drug doxorubicin (DOX) in immuno paramagnetic thermosensitive liposomes. DOX is the most common chemotherapeutic agent for the treatment of a variety of carcinomas. However, the pure drug has high cytotoxicity and therefore requires a targeted and biocompatible delivery system. The introduction includes concepts, modalities, and functionalities of the project. First, a detailed description of the cell type (triple-negative breast cancer) is given. Furthermore, the importance of liposomal doxorubicin is explained and the current state of research is shown. The importance of modification to achieve thermosensitive properties and the procedure for co-encapsulation with Gd chelate to achieve paramagnetic properties is also discussed. In addition, the first part describes the surface modification with ADAM8 antibodies, which leads to improved targeting. The second part of the thesis covers the different materials and methods used in this paper. The production of the liposomes LipTS, LipTS-GD, LipTS-GD-CY, LipTS-GD-CY-MAB and the loading of DOX using an ammonium sulfate gradient method were described in detail. The results part deals with the physicochemical characterization using dynamic light scattering and laser Doppler velocimetry, which confirmed a uniform monodisperse distribution of the liposomes. These properties facilitate the approach of liposomes to target cancer cells. The influence of lipid composition of liposomes, co-encapsulation with Gd chelate and surface modification of liposomes was evaluated and described accordingly. The size and structure of the individual liposomal formulations were determined by atomic force microscopy and transmission electron microscopy. Morphological examination of the liposomes confirmed agreement with the sizes obtained by dynamic light scattering. Temperature-dependent AFM images showed an intact liposome structure at 37 °C, whereas heating by UHF-MRI led to a lipid film indicating the destruction of the lipid bilayer. Furthermore, TEM images showed the morphological properties of the liposomes and gave a more precise indication of how Gd-chelate accumulates within the liposomes. Liposomes with Gd-chelate showed well-separated vesicles, suggesting that Gd- chelate is deposited in the lipid bilayer of the liposomes. Gd was encapsulated in the hydrophilic core whereas chelate was extended into the lipid bilayer. By differential scanning calorimetry and drug release, the heat-sensitive functionality of the liposomes could be determined. Liposomes showed a beginning of phase transition temperature at about 38 °C, which can be achieved by UHF-MRI exposure. The maximum phase transition temperature in the case of LipTS-GD and LipTS-GD-CY-MAB was 42 °C and 40 °C, respectively. A proof of concept study for the thermosensitive properties of liposomes and a time-dependent DOX release profile in hyperthermia was performed. Gd-chelate is encapsulated in both LipTS-GD and LipTS-GD-CY-MAB and led to paramagnetic properties of the liposomes. This facilitates imaging mediated DOX delivery and diagnosis of the solid tumor and metastatic cells. The change in relaxation rate R1 of liposomes was quantified before and after heating above Tm (T> Tm). The relaxivity of the liposomes was obtained from the adapted slope of the relaxation rate against the Gd concentration. Remarkably, the relaxation rate and relaxivity increased after heating the liposomes above Tm (T> Tm), suggesting that the liposomes opened, released Gd chelate, and the exchange of water molecules became faster and more practicable. Toxicity studies describe the different mechanisms for induced DOX toxicity. The increased cytotoxic effect at elevated temperatures showed that the induced toxicity is thermally dependent, i.e. DOX was released from the liposomes. The high viability of the cells at 37 °C indicates that the liposomes were intact at normal physiological temperatures. Under UHF-MRI treatment, cell toxicity due to elevated temperature was observed. The cellular uptake of liposomes under UHF-MRI was followed by a confocal laser scanning microscope. An increase in fluorescence intensity was observed after UHF-MRI exposure. The study of the uptake pathway showed that the majority of liposomes were mainly uptake by clathrin-mediated endocytosis. In addition, the liposomes were modified with anti-ADAM8 antibodies (MAB 1031) to allow targeted delivery. The cellular binding capabilities of surface-modified and non-modified liposomes were tested on cells that had ADAM8 overexpression and on ADAM8 knockdown cells. Surface-modified liposomes showed a significant increase in binding ability, indicating significant targeting against cells that overexpress ADAM8 on their surface. In addition, cells with knockdown ADAM8 could not bind a significant amount of modified liposomes. The biocompatibility of liposomes was assessed using a hemolysis test, which showed neglected hemolytic potential and an activated thromboplastin time (aPTT), where liposomes showed minimal interference with blood clotting. Hemocompatibility studies may help to understand the correlation between in vitro and in vivo. The chorioallantois model was used in ovo to evaluate systematic biocompatibility in an alternative animal model. In the toxicity test, liposomes were injected intravenously into the chicken embryo. The liposomes showed a neglectable harmful effect on embryo survival. While free DOX has a detrimental effect on the survival of chicken embryos, this confirms the safety profile of liposomes compared to free DOX. LipTS-GD-CY-MAB were injected into the vascular system of the chicken embryo on egg development day 11 and scanned under UHF-MRI to evaluate the magnetic properties of the liposomes in a biological system with T2-weighted images (3D). The liposomal formulation had distinct magnetic properties under UHF MRI and the chick survived the scan. In summary, immunomagnetic heat-sensitive liposomes are a novel drug for the treatment of TNBC. It is used both for the diagnosis and therapy of solid and metastasizing tumors without side effects on the neighboring tissue. Furthermore, a tumor in the CAM model will be established. Thereafter, the selective targeting of the liposomes will be visualized and quantitated using fluorescence and UHF-MRI. Liposomes are yet to be tested on mice as a xenograft triple-negative breast cancer model, in which further investigation on the effect of DOX-LipTS-GD-CY-MAB is evaluated. On one hand, the liposomes will be evaluated regarding their targetability and their selective binding. On the other hand, the triggered release of DOX from the liposomes after UHF-MRI exposure will be quantitated, as well as evaluate the DOX-Liposomes therapeutic effect on the tumor

NH2+ implantations induced superior hemocompatibility of carbon nanotubes

NH(2)(+) implantation was performed on multiwalled carbon nanotubes (MWCNTs) prepared by chemical vapor deposition. The hemocompatibility of MWCNTs and NH(2)(+)-implanted MWCNTs was evaluated based on in vitro hemolysis, platelet adhesion, and kinetic-clotting tests. Compared with MWCNTs, NH(2)(+)-implanted MWCNTs displayed more perfect platelets and red blood cells in morphology, lower platelet adhesion rate, lower hemolytic rate, and longer kinetic blood-clotting time. NH(2)(+)-implanted MWCNTs with higher fluency of 1 × 10(16) ions/cm(2) led to the best thromboresistance, hence desired hemocompatibility. Fourier transfer infrared and X-ray photoelectron spectroscopy analyses showed that NH(2)(+) implantation caused the cleavage of some pendants and the formation of some new N-containing functional groups. These results were responsible for the enhanced hemocompatibility of NH(2)(+)-implanted MWCNTs